Technical Abstract: The overuse of petrochemical-based synthetic fertilizers has caused detrimental effects to soil, water supplies, foods, and animal health. This, in addition to increased awareness of organic farming, has generated considerable interest in the evaluation of renewable biofertilizers. In order to study these effects, research was conducted to 1) evaluate and optimize a solid phase extraction procedure to extract three plant hormones, indole-3-acetic aicd (IAA), gibberellic acid (GA3), and abscisic acid (ABA), from two model biofertilizers produced from coconut and pineapple peels; 2) develop a HPLC analysis procedure for the simultaneous separation and quantification of the three plant hormones (IAA, GA3, and ABA); and 3) evaluate the changes in the levels of the three plant hormones at four different fermentation time periods while varying the number of general bacteria, lactic acid bacteria, and yeast. An optimized procedure for sample preparation, separation, and simultaneous analysis of three plant hormones produced in liquid biofertilizers was developed. The method involved sample cleanup using a sep-pack Oasis®MAX cartridge containing mixed-mode anion-exchange and reverse phase sorbents that provided optimum recovery of 85.6%, 91.9%, and 94.3%, respectively, for the three hormones IAA, GA3, and ABA. Baseline separation of the three hormones was achieved using a mobile phase consisting of 1% acetic acid and acetonitrile (75:25, v/v) at pH 4.0. The amounts of hormones produced in the liquid biofertilizers were influenced by fruit type, fermentation time, and total number of general bacteria, lactic acid bacteria, and yeast. The quantities of the three plant hormones produced during fermentation correlated well with the total number of microorganisms present in the liquid biofertilizers. This procedure allows quantifying the three plant hormones in their natural states without any prior derivatization step. This method can be extended to determine the quantity of the three hormones in other matrices. This assay procedure will aid in the development of liquid biofertilizers, a valuable alternative fertilizers to promote the plant growth. This process will help farmers reduce both production costs and pollution problems.